Method for the preparation of 21-hydroxy-6,19-oxidoprogesterone (21oh-6op)

ABSTRACT

The present invention relates to a novel method for preparing 21-hydroxy-6,19-oxidopro-gesterone (21OH-6OP) and/or its 21-acetate, 21-propionate, 21-hemisuccinate, 21-phosphate, 21-oleate derivatives. 21OH-6OP and its ester derivatives are antiglucocorticoids for the treatment or prophylaxis of diseases associated to an excess of glucocorticoids, in particular for treating Cushing&#39;s syndrome, iatrogenic hypercortisolism or depression.

FIELD OF THE INVENTION

[0001] The present invention relates to a novel method of preparing21-hydroxy-6,19-oxidopro-gesterone (21OH-6OP) and/or its 21-acetate,21-propionate, 21-hemisuccinate, 21-phosphate and 21-oleate derivatives.210H-60P and its esters are antiglucocorticoids for the treatment orprophylaxis of diseases associated with a glucocorticoid imbalance, inparticular for treating Cushing's syndrome or depression.

BACKGROUND OF THE INVENTION

[0002] Corticosteroides are steroid hormones related structurally tocholesterol. These hormones are synthesized in the adrenal cortex andinclude the glucocorticoids (e.g. cortisol), the mineralocorticoids (e.galdosterone) as well as weak androgens and estrogens. The adrenalfunction, like that of the thyroid gland, is under the control of thehypothalamus (HPT) and the pituitary (PIT). When cortisol (thenaturally-occuring glucocorticoid) levels drop below a setpoint, thehypothalamus releases CRH (corticotropin releasing hormone) whichstimulates adrenocorticotropic hormone (ACTH) release from thepituitary. ACTH is a tropic hormone which stimulates

[0003] the synthesis and secretion of cortisol (it has minimal effectson aldosterone synthesis/secretion), and

[0004] the growth of the adrenal gland. When cortisol levels increase,this shuts off CRH and ACTH secretion (cf. FIG. 1).

[0005] Cortisol is characterized by its properties related to thebiosynthesis and metabolism of glucose and propeties related tonon-specific as well as specific immunity. Due to their effects on theglucose metabolism, cortisol and natural or synthetic analogues thereofare usually named glucocorticoids. They bind to the glucocorticoidreceptor (GR).

[0006] The glucocorticoid receptor is a member of a protein super familyof closely related intracellular receptors which function asligand-activated transcription factors. Other members of this superfamily are the mineralocorticoid receptor (MR) and the progesteronereceptor (PR). MR and GR have shown to be highly homologous, thusnatural and even synthetic steroids exhibit cross-reaction between thesereceptors. With respect to PR, its natural ligand progesterone alsocross-reacts with MR and GR.

[0007] Cushing's syndrome is a disorder resulting from increasedadrenocortical secretion of cortisol. Hyperfunction of the adrenalcortex may be ACTH-dependent or it may be independent of ACTHregulation, e.g. production of cortisol by an adrenocortical adenoma orcarcinoma. The administration of supraphysiologic quantities ofexogenous cortisol or related synthetic analogs suppressesadrenocortical function and mimics ACTH-independent glucocorticoidhyperfunction. ACTH-dependent hyperfunction of the adrenal cortex may bedue to hypersecretion of ACTH by the pituitary, secretion of ACTH by anonpituitary tumor such as small cell carcinoma of the lung (the ectopicACTH syndrome), or administration of exogenous ACTH. While the term“Cushing's syndrome” has been applied to the clinical picture resultingfrom cortisol excess regardless of the cause, hyperfunction of theadrenal cortex resulting from pituitary ACTH excess has frequently beenreferred to as Cushing's disease, implying a particular physiologicabnormality. Patients with Cushing's disease may have a basophilicadenoma of the pituitary or a chromophobe adenoma. Microadenomas canusually be visualized by CT or, preferably, MRI scan, using ahigh-resolution technique augmented by gadolinium. Some micro-adenomasare difficult to visualize even with these modalities. In some cases, nohistological abnormality is found in the pituitary despite clearevidence of ACTH overproduction.

[0008] Reference to Cushing's syndrome is herein intended to mean theclinical picture resulting from cortisol excess regardless of the cause,which may be also iatrogenic, both by the injection of ACTH or by thedirect administration of cortisol or synthetic analogs such asprednisone, prednisolone, dexamethasone or others that are widely usedin various types of diseases including alergic, asthmatic, inflammatoryor immunologic. Cushing's syndrome includes in addition adrenal tumourssecreting corticoids, ectopic ACTH production and Cushing's disease.

[0009] Clinical manifestations include rounded “moon” faces with aplethoric appearance. There is truncal obesity with prominentsupraclavicular and dorsal cervical fat pads (“buffalo hump”); thedistal extremities and fingers are usually quite slender. Muscle wastingand weakness are present. The skin is thin and atrophic, with poor woundhealing and easy bruising. Purple striae may appear on the abdomen.Hypertension, renal calculi, osteo-porosis, glucose intolerance, reducedresistance to infection, and psychiatric disturbances are common.Cessation of linear growth is characteristic in children. Femalesusually have menstrual irregularities. An increased production ofandrogens, in addition to cortisol, may lead to hypertichosis, temporalbalding, and other signs of virilism in the female.

[0010] Although development of antihormonal agents related to theestrogen and androgen receptors has been successful, the search forselective anti-corticoids is more restricted.

[0011] Known agents suppressing the synthesis of steroid hormones atvarious levels (i.e. inhibitors of enzymes which catalyze various stagesof the synthesis of steroid hormones) are reviewed in J.SteroidBiochem., vol.5, p.501 (1974) and include the following:

[0012] a) derivatives of diphenyhnethane, e.g. amphenon B (whichsuppresses the synthesis of steroid hormones at stages 11-beta-, 17- and21- of hydroxylase);

[0013] b) derivatives of pyridine (SU-c series), e.g. metirapon (whichsuppresses synthesis at stage 11-beta of hydroxylase);

[0014] c) substituted alpha, alpha-glutaramides, e.g. aminoglutetimide(which impedes the synthesis of pregnenolone from cholesterol throughsuppression of 20-alpha-hydroxylase and C₂₀, C₂₂-liase;

[0015] d) steroid substances e.g. trilostan (3 beta-substitutedsteroid-3 beta hydroxy-5-androsten-17-one), which suppresses 3beta-desoxysteroidhydrogenase-5.4-isomerase (Steroids, vol.32, p.257).

[0016] e) steroids of the spironolactone family which are used asrapidly dissociating anti-Mineralocorticoids (PNAS USA 71(4) p.1431-1435 (1974).

[0017] f) a synthetic steroid described as an anti-Mineralocorticoids,ZK91587, showing specific binding properties for the kidney(Z.Naturforsch., 45b, p.711-715 (1990)) and hippocampus type I MR (LifeScience, 59, p.511-21 (1996)), but not for type II GR. It may thereforebe conveniently useful as a tool in the investigation of MR function intissues containing both receptor systems.

[0018] Agents that specifically suppress the interaction ofglucocorticoid hormones with hormone receptors are:

[0019] a) Mifepriston (11 β,17β)-11-[4-(Dimethylamino)phenyl]-17-hydroxy-17-(1-propynyl)estra-4,9-dien-3-one,which acts on receptors of glucocorticoid hormones to form a complexincapable of initiating mechanisms leading to glucocorticoid effect(Annals of New-York Academy of Science, vol. 761, p.5-28 (1995)).

[0020] b) non-steroid substances (J:Steroid Biochem., vol. 31, p.481-492(1988)) e.g. drotaverina hydrochloride (a derivative ofisoquinoline-1-(3.4-dietoxibenezilidene)-6.7-dietoxy-1,2,3,4-tetrahydrizoquinoline) or acetylsalicicacid (Moskovskaya Meditsina, 1990, “Receptor mechanisms of theglucocorticoid effect” by V. P. Golikov).

[0021] To-date, the only therapeutical application forantiglucocorticoids (e.g. Mifepristone) that has been attempted in aclinical setting is to treat inoperable cases of nonpituitary Cushing'ssyndrome. In the case of Mifepristone (both an anti-progesterone and ananti-glucocorticoid), high doses (up to 800 mg per day) are required.

[0022] Employing a systematic application of strategies to increaseactivity and decrease cross-reactivity and undesirable side effects,progress has been reported in the development of antihormonal agentswith greater potency and selectivity, especially in the antiestrogen andantiandrogen fields.

[0023] In EP-903'146 the synthetic steroid,21-hydroxy-6,19-oxidoprogesterone (21OH-6OP) of formula I,

[0024] is disclosed as a selective antiglucocorticoid which does notsubstantially cross-react with uterus-PR or kidney-MR. Said21-hydroxy-6,19-oxidoprogesterone antiglucocorticoid could be used inthe treatment of diseases associated with an excess of glucocorticoids,where an anti-glucocorticoid virtually lacking mineralocorticoid orglucocorticoid properties as well as affinity for MR or PR is desirable.

[0025] The synthesis of 21-hydroxy-6,19-oxidoprogesterone (1) and its21-acetate (2) was first accomplished by Deghenghi in 1966 asintermediate in a synthesis of 19-hydroxy-desoxy-corticosterone,starting from 21-hydroxypregnenolone diacetate. The procedure summarizedin Scheme 1 involves the use of a “hypoiodite type” reaction with leadtetraacetate (Pb(AcO)₄). Compound 1 was neither isolated norcharacterized, but acetylated in situ to acetate 2. Overall yield ofpartially purified 2 was only 8.3%. Further to the low yield, thismethod of preparation is generally perceived as being difficult toreproduce notably due to the formation of the chlorohydrin.

[0026] In a more recent synthesis of 19-hydroxydeoxycorticosterone, Kirkand Yeoh (J. Chem. Soc. Perkin Trans. I, 2945 (1983)) prepared acetate 2as an intermediate. This procedure starting from pregnenolone acetate isdepicted in Scheme 2. Although full details of the first 3 steps are notgiven in the experimental section of their publication, according to theliterature cited the yield for these steps may be estimated as ca.34-37% giving an overall yield of acetate 2 of ca. 12% from pregnenoloneacetate.

[0027] According to a further method, depicted in Scheme 3, the remotefunctionalization reaction with Pb(AcO)₄ and iodine under thermal orphotochemical conditions is replaced with the more reproducible and“milder” HgO/iodine system under photochemical conditions and the21-hydroxylation step is carried out with a hypervalent iodine compound(see A. S. Veleiro, M. V. Nevado, M. C. Monteserin and G. Burton,Steroids, 60, 268-272 (1995); M. Akhtar and D. H. R. Barton, J. Am.Chem. Soc., 86, 1528-1534 (1964); R. M. Moriarty, L. S. John and P. C.Du, J. Chem. Soc. Chem. Commun., 641-642 (1981).

[0028] The above procedure starts also from pregnenolone acetate whichis first converted into 21-hydroxypregnenolone diacetate. In the bestcase, overall yield of 1 is ca. 26%; however this product, althoughapparently pure according to TLC and NMR, could not be crystallized, andthus required additional purification steps by chromatography; yield ofpure crystalline 1 is about 13% (19% from 21-hydroxypregnenolonediacetate). Although the acetate 2 is an intermediate in this syntheticroute, complete purification could only be achieved after deacetylation(i.e. on compound 1). Furthermore, although this procedure allows thesynthesis of small amounts of 1 for the initial biological tests carriedout in 1996, attempts to scale up the procedure failed. Although, thecurrently known methods provide 21-hydroxy-6,19-oxidoprogesterone (1),they consistently give poorer yields as well as byproducts which aredifficult to eliminate.

DESCRIPTION OF THE INVENTION

[0029] It is an object of the invention to provide a new method forpreparing 21-hydroxy-6,19-oxidopro-gesterone (21OH-6OP) and/or itsesters.

[0030] It is a further object of the invention to provide21-hydroxy-6,19-oxidoprogesterone and/or its esters.

[0031] In a first aspect, the present invention provides a new method ofpreparing 21-hydroxy-6,19-oxidoprogesterone (1) or its esters, forexample carboxylate, phosphate or sulfate esters. In preferredembodiments, the invention provides a new method for preparing 21 OH-6OPand its 21-acetate, 21-propionate, 21-hemisuccinate, 21-phosphate and21-oleate derivatives. The method of the invention avoids essentiallyall of the hitherto known inconveniences. The method of the inventionproduces 21-hydroxy-6,19-oxidoprogesterone (1) in good yield, having anacceptable degree of purity, and avoids heavy metals as reagent. Themethod of the invention is suitable for large scale industrialpreparation of 21-hydroxy-6,19-oxido-progesterone (1) and its esters.

[0032] The objects of the invention are met according to the main claim.Preferred embodiments are set out within the dependent claims which areincorporated herewith.

[0033] The novel synthetic procedure according to the present inventioncomprises or consists of the following consecutive basic steps:

[0034] a) Providing 21-acetoxypregnenolone (3) (which is actually acommercial product);

[0035] b) Transforming the C-3 hydroxy group of 21-acetoxypregnenoloneinto a labile ester, preferably a formate ester;

[0036] c) Obtaining the bromohydrin product from the protected21-acetoxypregnenolone, said bromohydrin resulting from the addition ofa bromine and a hydroxy group onto the double bond of position of C₅-C₆;

[0037] d) Performing an intramolecular cyclisation with the C19 atomwith the “Suarez-reagent” (diacetoxyiodobenzene; see Annas et al., J.Chem.Soc. Perkin I, 405 (1989)) and iodine under irradiation, thusobtaining the 6,19 oxido-bridge within the scaffold;

[0038] e) Performing a selective hydrolysis, preferably withHCl/MeOH/dichloromethane, followed by an oxidation, preferably withpyridinium chlorochromate (PCC), thus obtaining a bromoketone;

[0039] f) Performing a hydrolysis of the bromoketone resulting from stepe) to obtain 21-hydroxy-6,19-oxidoprogesterone (1), and optionally

[0040] g) Acylating 21-hydroxy-6,19-oxidoprogesterone (1), to afford the21-ester (the acetate is shown as 2);

[0041] Preferred esters are C1-18 acyl esters (optionally substitutedwith COOH, 1 or 2 times) and phosphate esters. Acyl esters may beobtained by reacting 21-hydroxy-6,19-oxidoprogesterone (1) with anorganic acid, in the presence of a coupling agent (for exampleN,N′-dicyclohexylcarbodiimide), or with an activated organic ester (forexample, a nitrophenol ester), or with an acyl halide (for example, anacyl chloride), or with an acyl anhydride. Phosphate esters may beobtained by reacting 21-hydroxy-6,19-oxidoprogesterone with aphosphorylating agent (for example phosphorus oxychloride, followed bybasic hydrolysis).

[0042] The 21-propionate or 21-hemisuccinate, and 21-oleate andderivatives are obtained by esterifying21-hydroxy-6,19-oxidoprogesterone (1) of step f) with propionic acid,succinic or oleic acid, their anhydrides, activated esters or acylchlorides.

[0043] A preferred synthetic procedure according to the presentinvention is depicted in Scheme 4.

[0044] According to the preferred method illustrated in Scheme 4, aformate group is introduced as protecting group for position 3. Formateshave the advantage that they may be hydrolized under relatively mildacid conditions (HCl/MeOH-dichloromethane) in which primary acetates andeven α-acetoxy-ketones are stable. The introduction of a formate moietyis preferably carried out under very mild conditions using preferablymixed acetic-formic anhydride (prepared in situ from formic acid andacetic anhydride). The bromo-hydrin formation, is carried out withN-bromoacetamide in THF yielding 80% of the desired bromohydrin 5 andabout 20% of the 5α-HO-6β-Br isomer. The mixture may be separated bycrystallization or Thin Layer Chromatography. For the intramolecularcyclisation involving C-₁₉, the “Suarez reagent” (diacet-oxyiodobenzene(DAIB)) and iodine is employed as the “hypoiodite” as generating systemunder suitable irradiation—preferably with a standard tungsten lamp.

[0045] Quite surprisingly, it turns out that upon performing theintramolecular cyclisation step in di-chloromethane (CH₂Cl₂), thereaction is not only rapid (complete conversion in less than 1 hour) andclean, but also excellent yields could be obtained.

[0046] Temperature control (25° C.) of the cyclisation step, achieved byusing a glass jacketed reactor with water circulation, increases theyield and diminishes excessive oxidation by-products. Furthermore,epoxide formation is eliminated, the isomeric bromohydrin is inert tothe reagent and could be easily separated (together with the iodobenzeneside product) from the desired bromoether 6 by vacuum filtration througha silica gel column (VFC).

[0047] The selective hydrolysis of the formate ester, is preferablyperformed with HCl in MeOH-dichloromethane, followed by an oxidation,preferably with PCC, thus affording the bromoketone 8. This productpartially eliminates HBr to give the unsaturated ketone (2) whensubjected to VFC (very short silica gel column) purification, howeverthis poses no problem as the unsaturated product is the desired productof the following step, in which the treatment with a base deacetylatesposition 21 and completes the elimination of the C-5 bromine thus givingthe desired pro-duct (1).

[0048] Purification of 1 may be achieved with a fast VFC through a shortsilica gel column to afford chromatographically pure product in over 31%yield. This may be crystallized from absolute ethanol to yieldcrystalline 1 in 27% yield.

[0049] The 21-acetate (2) is prepared by standard acetylation of 1 withacetic anhydride in pyri-dine. The acetate is recrystallized fromabsolute ethanol.

[0050] The novel synthesis according to the present invention has thefollowing advantages:

[0051] a) It eliminates the need of the selective acetylation step orany need to differentiate the hydroxyl groups at C-21 and C-3.

[0052] b) It modifies the hypoiodite reaction so that no heavy metalsare required (lead, silver or mercury based reagents) and it proceedsunder homogeneous conditions, thus allowing a scale up to multigram andeventually kilogram amounts.

[0053] c) It reduces the secondary products formed at different stages(specially 5,6-epoxide formation) so as to minimize the purificationsteps, and specially to avoid or minimize the need of columnchromatography on silica gel, as bromoethers, compound 1 and 2 areintrinsically unstable under these conditions.

[0054] d) It reduces reaction volumes throughout to allow scale up.

[0055] Starting from compound 1, synthesis of acyl esters, such as21-propionate (2a), and 21-hemisuccinate (2b) and 21-oleate(2c)-6,19-oxidoprogesterone derivatives can be performed as illustratedin Scheme 5.

[0056] Also shown in Scheme 5 is the preparation of the 21-phosphatederivative (2d).

[0057] The compounds synthesised according to the present invention arefor use as a medicament, alone or in combination with pharmaceuticallyacceptable carriers and/or excipients. Such medicament is notablysuitable in the manufacture of a medicament for the treatment orprophylaxis of diseases associated with an excess of glucocorticoids,e.g. for the treatment of Cushing's syndrome, iatrogenichypercortisolism or depression.

[0058] The invention will now be described, by way of illustration only,with reference to the following examples:

EXAMPLES

[0059] Materials and Methods

[0060] Melting points were taken on a Fisher-Johns apparatus and areuncorrected. IR spectra were recorded in thin films using KBr disks on aNicolet Magna IR 550 FT-IR spectrometer. ¹H and ¹³C NMR spectra weremeasured in Bruker AC-200 or AM-500 NMR spectrometers indeu-teriochloroform (using TMS as internal standard). The J values aregiven in Hz. Spectra were assigned by analysis of the DEPT, COSY 45 andHETCOSY spectra and by comparison with those of progesterone.

[0061] The electron impact mass spectra (EI) were measured in a VG Trio2 mass spectrometer at 70 eV by direct inlet. FAB mass spectra andelectron impact high resolution mass spectra (HRMS) were obtained in aVG ZAB BEQQ mass spectrometer. All solvents used were reagent grade.Solvents were evaporated at ca. 45° C. under vacuum. Zinc dust wasactivated by suspending in 1M HCl, washing with water, absolute ethanoland diethyl ether and drying 2 h at 120° C. The homogeneity of allcompounds was confirmed by thin layer chromatography.

[0062] Large Scale Synthesis of 21-hydroxy-6,19-oxidoprogesterone (1)

[0063] Structures of starting material, intermediates and final products(1) and (2):

[0064] 3β-Formyloxy-21-acetyloxy-5-pregnen-20-one (4)

[0065] Acetic anhydride (13.4 ml) is added dropwise to formic acid (6.6ml) at 0° C., the solution is heated at 50° C. for 15 min and cooledrapidly to 0° C. The resulting acetoformic anhydride solution is addeddropwise to a stirred suspension of 21-acetoxypregnenolone (3, 8.0 g) indry pyridine (20.8 ml) at 0° C., and stirring is 15 continued at thattemperature for 2 h. The reaction product is poured over cold saturatedaqueous sodium bicarbonate solution, filtered and the solid is washedwith saturated aqueous sodium bicarbonate solution, water and 1N HCl andwater (until neutral) rendering formate 4 (8.0 g); ¹H NMR (200.13 MHz)δ_(H) 0.70 (3H, s, 13-CH₃), 1.02 (3H, s, 10-CH₃), 2.16 (3H, s,21-CH₃CO), 2.53 (1H, t, J=8.0 Hz, 17-H), 4.50 (1H, d, J=17.0 Hz, 21a-H), 4.70 (1H, d, J=17.0 Hz, 21b-H), 5.32 (1H, m, 3-H), 5.38 (1H, d,J=3.0 Hz, 6-H), 8.02 (1H, s, HCOO).

[0066] 3β-Formyloxy-5α-bromo-6β-hydroxy-21-acetyloxypregnan-20-one (5)

[0067] Formate 4 (8.0 g), is dissolved in diethyl ether (100 ml) and THF(37.2 ml) and cooled to 10° C. To the stirred solution at 10-15°C.—which protected from light—7.5% perchloric acid (11.88 ml) is added,followed by N-bromoacetamide (4.75 g) in 8 portions over a 25 minperiod. Stirring is continued for 45 min at 25° C. and the reaction isstopped by addition of 10% aqueous sodium thiosulfate solution untilcomplete decoloration. The reaction mixture is then extracted withdichloromethane/methanol 10:1 and the organic layer, is washed withwater, dried with anhydrous sodium sulfate and the solvent is evaporatedto afford bromohydrin 5 (10.4 g, containing about 20% of the5α-hydroxy-6β-bromo isomer as determined by ¹H NMR).

[0068] 3β-Formyloxy-5α-bromo-21-acetyloxy-6,19-oxidopregnan-20-one (6)

[0069] Nitrogen is bubbled for 5 min through a solution of bromohydrincompound 5 (10.4 g, containing about 20% of the 5α-hydroxy-6μ-bromoisomer) in freshely distilled dichloro-methane (723 ml) contained in a 1liter glass vessel fitted with an external cooling jacket withcirculating water at 25° C. and magnetic stirrer.

[0070] Diacetoxyiodobenzene (Suarez reagent, 7.66 g) and iodine (5.46 g)are successively added with stirring. The vessel is exposed to two 300Watt tungsten lamps (5000 lm each) and vigorous stirring is continuedfor 1 h at 25° C. Irradiation is turned off and a saturated aqueoussolution of sodium thiosulfate is added until complete decoloration. Theorganic layer is separated, dried with anhydrous sodium sulfate and thesolvent evaporated. The resulting solid is dissolved in dichloro-methane(8 ml) and applied to a silicagel G-60 column (12 cm diameter×8 cmheight) previously flushed with hexane; successive elution (applyingvacuum to the outlet) with hexane-ethyl acetate 9:1 (1100 ml), 8:2 (700ml), 7:3 (700 ml) and 6:4 (600 ml) affords 31×100 ml fractions.Fractions are analyzed by TLC and those containing bromoether 6 arepooled and evaporated to dryness to afford 6 (6.8 g). ¹H NMR (200.13MHz) δ_(H) 0.70 (3H, s, 13-CH₃), 2.16 (3H, s, 21-CH₃CO), 2.52 (1H, t,J=8.8 Hz, 17-H), 3.73 (1H, d, J=8.4 Hz, 19a-H), 3.94 (1H, d, J=8.4 Hz,19b-H), 4.08 (1H, d, J=4.2 Hz, 6-H), 4.50 (1H, d, J=16.8 Hz, 21a-H),4.71 (1H, d, J=16.8 Hz, 21b-H), 5.34 (1H, m, 3-H), 8.02 (1H, s, HCOO).

[0071] 3β-Hydroxy-5α-bromo-21-acetyloxy-6,19-oxidopregnan-20-one (7)

[0072] A stirred solution of the bromoether 6 (6.8 g) obtained above, isdissolved in dichloro-methane (45.7 ml) and methanol (154.7 ml) and iscooled to 0° C. in an ice bath and water (10.9 ml) while conc. HCl (23.0ml) is added. After about 30 min of vigorous stirring at 0° C.(disappearance of the starting material is monitored by TLC) thereaction mixture is neutralized with 20% aqueous sodium hydroxide andextracted with dichloromethane. The organic layer is dried withanhydrous sodium sulfate and the solvent evaporated to afford thealcohol compound 7 (6.5 g); ¹H NMR (200.13 MHz) δ_(H) 0.69 (3H, s,13-CH₃), 2.16 (3H, s, 21-CH₃CO), 2.52 (1H, t, J=8.5 Hz, 17-H), 3.62 (1H,d, J=8.5 Hz, 19a-H), 3.92 (1H, d, J=8.5 Hz, 19bH), 4.07 (1H, d, J=4.0Hz, 6-H), 4.15 (1H, m, 3-H), 4.51 (1H, d, J=17.0 Hz, 21aH), 4.70 (1H, d,J=17.0 Hz, 21b-H).

[0073] 5α-Bromo-21-acetyloxy-6,19-oxidopregnane-3,20-dione (8)

[0074] A suspension of pyridinium chlorochromate (12.1 g), bariumcarbonate (5.0 g) and 3 Å molecular sieves (9.60 g), in drydichloromethane (480 ml) is stirred under nitrogen for 10 min. To theresultant orange slurry a solution of bromoether 7 (6.5 g) obtainedabove in dry dichloromethane (324 ml) is added and stirring is continuedfor about 90 min, until the starting material (TLC) has disappeared. Thereaction mixture is percolated through a short silicagel G 60 column (12cm diameter×8 cm height) washed with diethyl ether (2×150 ml) andhexane-ethyl acetate 1:2 (3×150 ml). Fractions containing the productare pooled and evaporated to dryness affording 5.5 g of ketone 8(containing about 10% of Δ⁴-3-ketone (2); ¹H NMR (200.13 MHz) δ_(H) 0.70(3H, s, 13-CH₃), 2.16 (3H, s, 21CH₃CO), 2.51 (1H, t, J=8.5 Hz, 17-H),2.85 (1H, d, J=16.0 Hz, 4a-H), 3.40 (1H, d, J=16.0 Hz, 4b-H), 3.90 (1H,d, J=9.0 Hz, 19a-H), 4.07 (1H, d, J=4.0 Hz, 6H), 4.15 (1H, d, J=9.0 Hz,19b-H), 4.50 (1H, d, J=17.0 Hz, 21a-H), 4.71 (1H, d, J=17.0 Hz, 21b-H).

[0075] 21-Hydroxy-6,19-oxido-4-pregnene-3,20-dione (1)

[0076] The ketone 8 (5.5 g) obtained from the preceding step isdissolved in methanol (263.8 ml) and dichloromethane (13.2 ml). To thestirred solution 14% methanolic KOH (53.4 ml) is added and stirring iscontinued at room temperature for about 15 min, until the startingmaterial (TLC) has disappeared. The reaction mixture is neutralized with1N HCl and extracted with dichloromethane. The organic layer is driedwith anhydrous sodium sulfate and the solvent is evaporated thusyielding crude 21-hydroxy-6,19-oxidoprogesterone (1, 4.6 g). The solidis dissolved in dichloromethane (5 ml) and sent through to a silicagelG-60 column (8.5 cm diameter×5 cm height) previously flushed withhexane-ethyl acetate 7:3; successive elution (applying vacuum to theoutlet) with hexane-ethyl acetate 6:4 (1350 ml) and 1:1 (900 ml) affords30 fractions. The fractions are analyzed by TLC and those containing 1are pooled and evaporated to dryness to afford21-hydroxy-6,19-oxidoprogesterone (1, 2.3 g). ¹H NMR (200.13 MHz) δ^(H)0.74 (3H, s, 13-CH₃), 2.45 (1H, t, J=8.5 Hz, 17-H), 3.51 (1H, d, J-=8.8Hz, 19a-H), 4.18 (3H, s, 21-CH₃), 4.20 (1H, d, J=8.8 Hz, 19b-H), 4.69(1H, d, J=5.0 Hz, 6-H), 5.82 (1H, s, 4-H).

[0077] Re-crystallization from absolute ethanol affords a first crop ofcrystalline 1 (1.27 g), mp 165-166° C. The mother liquor is concentratedto yield a second crop of 1 (0.68 g).

[0078] 21-Acetyloxy-6,19-oxido-4-pregnene-3,20-dione (2)

[0079] Crude 21-hydroxy-6,19-oxidoprogesterone (1, 2 g beforechromatographic purification) was dissolved in dry pyridine (15.6 ml)and acetic anhydride (15.6 ml) added. The solution was stirred for 90min at 25° C., poured over 1M HCl and filtered (alternatively the solidmay be extracted with dichloromethane). The solid was washed with water(until neutral), dried, dissolved in dichlorometane (2 ml) and appliedto a silicagel G-60 column (7 cm diameter×5 cm height) previouslyflushed with hexane-ethyl acetate 7:3; succesive elution (applyingvacuum to the outlet) with hexane-ethyl acetate 7:3 (700 ml) and 6:4(700 ml) afforded 20 fractions. Fractions were analyzed by TLC and thosecontaining 2 pooled and evaporated to dryness to afford21-acetyloxy-6,19-oxidoprogesterone (2, 1.12 g). Recrystallization fromabsolute ethanol (with drops of methanol) afforded crystaline 2 (0.72g), mp 190-191° C. ¹H NMR (200.13 MHz) δ_(H) 0.76 (3H, s, 13-CH₃), 2.17(3H, s, 21-CH₃CO), 2.51 (1H, t, J=8.5 Hz, 17-H), 3.51 (1H, d, J=8.2 Hz,19a-H), 4.20 (1H, d, J=8.2 Hz, 19b-H), 4.50 (1H, d, J=16.7 Hz, 21a-H),4.69 (1H, d, J=5.0 Hz, 6-H), 4.72 (1H, d, J=16.7 Hz, 21b-H), 5.82 (1H,s, 4-H).

[0080] 21-Propanoyloxy-6,19-oxido-4-pregnene-3,20-dione (2a)

[0081] Crude 21-hydroxy-6,19-oxidoprogesterone (1, 0.2 g beforechromatographic purification) was dissolved in dry pyridine (0.28 ml)and propanoic anhydride (0.2 ml) added. The solution was stirred for 1 hat 25° C., methanol was added to destroy excess anhydride and thesolution concentrated in vacuo. The residue was diluted withdichloromethane, washed with 1M HCl and water, and evaporated to dryness(0.233 g). Purification as above afforded21-propanoyloxy-6,19-oxidoprogesterone (2a, 0.116 g). ¹H NMR (200.13MHz) δ_(H) 0.76 (3H, s, 13CH₃), 1.18 (3H, t, J=7.6 Hz, 21-CH₃CH₂CO),2.46 (2H, q, J=7.6 Hz, 21CH₃CH₂CO), 2.51 (1H, t, J=8.0 Hz, 17-H), 3.51(1H, d, J=8.2 Hz, 19a-H), 4.20 (1H, d, J=8.2 Hz, 19b-H), 4.50 (1H, d,J=16.8 Hz, 21a-H), 4.70 (1H, d, J=5.0 Hz, 6-H), 4.73 (1H, d, J=16.8 Hz,21b-H), 5.82 (1H, s, 4-H). EIMS m/z 400 (17) [M]+, 342 (5), 313 (24),285 (23), 267 (10), 57 (100).

[0082] 21-Succinoyloxy-6,19-oxido-4-pregnene-3,20-dione (2b)

[0083] 21-hydroxy-6,19-oxidoprogesterone (1, 0.75 g) was dissolved indry dichloromethane (37.5 ml) and pyridine (1.9 ml) and succinicanhydride (0.75 g) added. The solution was stirred for 4 h at 25° C., asecond portion of succinic anhydride (0.375 g) added and stirringcontinued for 6 h (until disappearance of starting material). Thereaction mixture was concentrated, and the residue extracted withdiethyl ether. The ethereal solution was washed with 1M HCl andextracted with aqueous 10% sodium carbonate. The aqueous layer wasacidified with conc. HCl to pH=3 and extracted with dichloromethane.Washing with water and evaporation to dryness afforded crude21-succinoyloxy-6,19-oxidoprogesterone (2b, 0.823 g). The solid wasdissolved in dichloromethane (1 ml) and applied to a silicagel G-60column (6 cm diameter×4 cm height) previously flushed with hexane-ethylacetate 2:8; succesive elution (applying vacuum to the outlet) withhexane-ethyl acetate 2:8 (200 ml) and ethyl acetate (500 ml) afforded 14fractions. Fractions were analyzed by TLC and those containing 2b pooledand evaporated to dryness to afford21-succinoyloxy-6,19-oxidoprogesterone (2b, 0.455 g). ¹H NMR (200.13MHz) δ_(H) 0.75 (3H, s, 13-CH₃), 2.52 (1H, t, J=8.0 Hz, 17-H), 2.75 (4H,m, 21-HCOOCH₂CH₂CO), 3.51 (1H, d, J=8.2 Hz, 19a-H), 4.20 (1H, d, J=8.2Hz, 19b-H), 4.54 (1H, d, J=16.9 Hz, 21a-H), 4.70 (1H, d, J=5.0 Hz, 6-H),4.75 (1H, d, J=16.9 Hz, 21b-H), 5.82 (1H, s, 4-H). EIMS m/z 444 (3)[M]+, 344 (24), 313 (55), 285 (44), 267 (17), 91 (60), 79 (54), 55(100).

[0084] 21-Oleoyloxy-6,19-oxido-4-pregnene-3, 20-dione (2c)

[0085] 21-hydroxy-6,19-oxidoprogesterone (1, 0.052 g) was dissolved indry dichloromethane (1 ml) and pyridine (0.12 ml) and oleoyl chloride(0.1 ml) added. The solution was stirred for 24 h at 25° C., dilutedwith dichloromethane, washed with 1M HCl and water, and evaporated todryness. Purification by preparative TLC afforded21-oleoyloxy-6,19-oxidoprogesterone (2c, 0.075 g). ¹H NMR (200.13 MHz)δ_(H) 0.76 (3H, s, 13-CH₃), 0.88 (3H, t, J=7.0 Hz, CH₃—CH—CO), 2.34 (2H,t, J=7.7 Hz, CH—CH₂CO), 2.50 (1H, t, J=8.5 Hz, 17H), 3.51 (1H, d, J=8.3Hz, 19a-H), 4.20 (1H, d, J=8.3 Hz, 19b-H), 4.49 (1H, d, J=16.9 Hz,21a-H), 4.70 (1H, d, J=4.5 Hz, 6-H), 4.73 (1H, d, J=16.9 Hz, 21b-H),5.34 (2H, m, CH—CH═CH—CH—CO), 5.82 (1H, s, 4-H).

[0086] 21-Phosphate-6,19-oxido-4-pregnene-3,20-dione (2d)

[0087] 21-hydroxy-6,19-oxido-4-pregnene-3,20-dione (1, 0.052 g) wasdissolved in dry dichloromethane (1 ml) and pyridine (0.12 ml), andphosphochloridic acid diallyl ester (0.024 g, 1 eq.) was added dropwiseat 0° C., over 30 minutes. The mixture was allowed to warm to roomtemperture and stirred overnight. The mixture was washed three timeswith 5% NaHCO₃, the organic layer was dried and evaporated in vacuowithout heating. To the resulting syrup was added a solution of NaOH(2,2 equivalents) in water (3 ml). The solution was brought slowly toreflux, and gently refluxed for 12 hours. Lyophilization yielded thecrude phosphate disodium salt, which was recrystallized from ethanol.Alternatively, the crude disodium salt may be dissolved in water, andthe pH adjusted with HCl, to cause precipitation of the free phosphate.

1. A method of preparing 21-hydroxy-6,19-oxidoprogesterone (1) or anacyl or phosphate derivative thereof.

the method comprising the steps a) providing 21-acetoxypregnenolone; b)protecting the C-3 hydroxy group of 21-acetoxypregnenolone throughformation of a labile ester; c) obtaining the bromohydrin productthereof, by addition of a bromine and a hydroxy group onto the doublebond of position of C₅-C₆ of the ester protected21-acetoxy-pregnenolone; d) performing an intramolecular cyclizationwith the C19 atom with the “Suarez-reagent” (diacetoxyiodobenzene) andiodine under irradiation, thus obtaining the 6,19 oxido-bridge withinthe scaffold; e) performing a selective hydrolysis, followed by anoxidation, to obtain a bromo-ketone; f) performing a hydrolysis of thebromoketone resulting from step e) to obtain21-hydroxy-6,19-oxidoprogesterone (1); and optionally g) reactingcompound (1) with an acylating agent or phosphorylating agent.
 2. Themethod according to claim 1, wherein the 21-acetate derivative isobtained by acetylating 21-hydroxy-6,19-oxidoprogesterone (1) of stepf).
 3. The method according to claim 1, wherein a C1-18 acyl derivativeis obtained by esterifying 21-hydroxy-6,19-oxidoprogesterone with aC1-18 acid, anhydride, active ester or acyl chloride.
 4. The methodaccording to claim 1, wherein a 21-propionate, 21-hemisuccinate,21-phosphate, 21-oleate derivative is obtained by esterifying21-hydroxy-6,19-oxidoprogesterone (1) of step f) with propionic acid,succinic acid, oleic acid or their anhydrides or active esters or acidchlorides.
 5. The method according to claim 1, wherein the 21-phosphatederivative is obtained by reacting oxidoprogesterone (1) of step (f)with a phosphorylating agent.
 6. The method according to any of thepreceding claims wherein for step b) a formate ester is afforded.
 7. Themethod according to any of the preceding claims, wherein theintramolecular cyclisation step d) is performed in dichloromethane assolvent.
 8. The method according to any of the preceding claims, whereinthe selective hydrolysis of step e) is performed with HCl inMeOH/dichloromethane as solvent.
 9. The method according to any of thepreceding claims, wherein the oxidation in step e) is performed withpyridinium chlorochromate (PCC).
 10. The method according to any of thepreceding claims, wherein the hydrolysis in step f) is performed withKOH in MeOH/dichloromethane as solvent.
 11. A method for preparing (6)

comprising the step of performing an intramolecular cyclisation betweenthe 6-OH and the C₂₁ of the molecule (5)


12. A method according to claim 11, wherein the intramolecularcyclisation is carried out by photoreaction with DIAB and I₂.
 13. Amethod according to claim 11 or 12, wherein the intramolecularcyclisation is carried out in CH₂Cl₂.